Steroid hormone regulation of nuclear proto-oncogenes.

The role of the nuclear proto-oncogenes as rapidly responding nuclear regulators in the cascade model for steroid hormone action is proposed. In this model, the nuclear proto-oncogenes respond within minutes to steroids and would code for regulatory proteins that in turn enter the nucleus to positively or negatively regulate "late" structural gene transcription and mRNA processing. The potential involvement of the nuclear matrix and one of its components, a receptor binding factor (RBF-1) for steroid receptors of these genes, is discussed. The nuclear proto-oncogenes, thus, may serve as important "early" regulatory genes and as excellent universal markers in all tissues in steroid hormone action. Proto-oncogenes are known to be regulatory genes since genetic alterations of amplification, mutations, chromosome translocation, and other rearrangements which result in overexpression, inactivation, and/or loss of regulatory control result in malignant transformation of the cell. It is now known that as many as six different proto-oncogenes must undergo alteration to induce the loss of cellular control of replication and/or for transformation to occur. The nuclear oncogenes, representing only one class of several of the known proto-oncogenes, code for transcription factors and enzymes involved in regulating these factors and other features of gene transcription. Possibly RNA processing is another function. Many of these genes and their responses to steroids are described in this chapter. The steroid receptors belong to a family which themselves represent transcription factors in that they reside in the nucleus and bind to specific DNA elements or other transcription factors to alter gene transcription and/or mRNA processing. The receptors for the various steroids are described as having a common structural motif and function using the above mentioned pathways. In the majority of cases cited, the steroid receptors mediate the rapid regulation of the nuclear proto-oncogene transcription. The role of palindromic SRE in or near these genes is to bind steroid hormone receptor dimers to regulate transcription. More powerful steroid response units (SRU), composed of two or more response elements for steroids or transcription factors, can act at great distances from the gene as enhancers for steroid- regulated transcription. What has become obvious from the studies of steroid effects on nuclear proto-oncogenes, is that, despite the tremendous scientific strides that have been made toward understanding gene regulation by steroids, we are still relatively naive.(ABSTRACT TRUNCATED AT 400 WORDS)

Nuclear matrix acceptor binding sites for steroid hormone receptors: a candidate nuclear matrix acceptor protein.

Steroid/nuclear-hormone receptors are ligand-activated transcription factors that have been localized to the nuclear matrix. The classic model of hormone action suggests that, following activation, these receptors bind to specific "steroid response elements" on the DNA, then interact with other factors in the transcription initiation complex. However, evidence demonstrates the existence of specific chromatin proteins that act as accessory factors by facilitating the binding of the steroid receptors to the DNA. One such protein, the "receptor binding factor (RBF)-1", has been purified and shown to confer specific, high-affinity binding of the progesterone receptor to the DNA. Interestingly, the RBF-1 is localized to the nuclear matrix. Further, the RBF-1 binds specifically to a sequence of the c-myc proto-oncogene that has the appearance of a nuclear matrix attached region (MAR). These results, and other findings reviewed here, suggest that the nuclear matrix is involved intimately in steroid hormone-regulated gene expression.

Mid-arm approach to basilic and cephalic vein cannulation using ultrasound guidance.

Central venous lines are often used when there is difficulty obtaining peripheral venous access. The basilic and cephalic veins in the mid-arm region, although difficult to see or palpate, can be imaged longitudinally and cannulated using real-time ultrasonography, providing an easy alternative. These techniques are described, with reports of four typical cases.

Open management of septic abdomen by Marlex mesh zipper.

Marlex mesh with zipper was used for abdominal closure in 5 of 147 patients with generalized peritonitis seen during a period of 2 years. Residual/recurrent intra-abdominal sepsis necessitating repeated explorations prompted use of this technique followed by frequent peritoneal lavages. Abdominal sepsis was successfully controlled in 4 of 5 patients, although we lost 3 of 5 patients due to multiple factors.

A nuclear matrix acceptor site for the progesterone receptor in the avian c-myc gene promoter.

It has been the goal of this project to determine the location, composition, and biological function of the nuclear acceptor sites (i.e., the nuclear binding sites) for the avian oviduct progesterone (Pg) receptor (PR). Many laboratories have demonstrated a native-(in vivo) like cell-free binding of steroid receptor complexes to specific acceptor sites in nuclei/chromatin in a variety of target tissue systems. These sites appear to involve protein-DNA complexes and some of these have been shown to reside in the nuclear matrix, including the chromatin acceptor sites for the avian oviduct PR. We have purified a nuclear matrix "acceptor protein" for the avian PR. termed receptor binding factor-1 (RBF-1), based on its ability to generate specific, high-affinity PR binding on avian genomic DNA. This 10 kD nuclear matrix protein was found to be unique with minimal homology to a couple of other proteins. Using immunohistochemical techniques and antibodies against the purified RBF-1, the RBF-1 was localized to the nuclei of many avian and rat tissues. Co-localizations of RBF-1 and PR in select cell types in the avian oviduct and rat reproductive organs were also reported. A tissue specificity was found with regard to RBF-1 concentrations. The full length cDNA to RBF-1 has been isolated and used to identify a 0.7 kb mRNA whose levels in various avian tissues reflect the protein levels. Genomic sequences of RBF-1 have been isolated and characterized. Preliminary studies indicate that the over-expression of the RBF in human MCF-7 cells results in an inhibition of the c-myc gene promoter activity which is further inhibited by steroid hormone treatments of the cells. Past studies in our laboratory demonstrated that the c-myc mRNA steady state levels are rapidly (approximately 15 min) reduced by Pg and glucocorticoids in the avian oviduct. Further, partially purified fractions of RBF-1 were shown to generate specific PR binding sites only on the genomic DNAs of certain animal species and on the c-myc gene, but not ovalbumin gene. Using Southwestern blot analyses, the purified RBF-1 was shown to bind specifically to sequences in the 5' end of c-myc, c-jun proto-oncogenes but not to genomic sequences of the ovalbumin gene. A specific DNA binding element in the promoter region of the c-myc proto-oncogene has been identified as AT-rich domain of homopurine/pyrimidine stretches flanked by GC-rich sequences. Southern blot analyses using 200 bp matrix DNA fragments protected by the nuclear matrix structure indicate that the matrix is attached on either side of the RBF-1 binding element. A model is described for a nuclear matrix acceptor site attached to the c-myc promoter which may mediate the Pg-induced down-regulation of the c-myc gene expression.

Interactions of the nuclear matrix-associated steroid receptor binding factor with its DNA binding element in the c-myc gene promoter.

Steroid receptor binding factor (RBF) was originally isolated from avian oviduct nuclear matrix. When bound to avian genomic DNA, RBF generates saturable high-affinity binding sites for the avian progesterone receptor (PR). Recent studies have shown that RBF binds to a 54 bp element in the 5'-flanking region of the progesterone-regulated avian c-myc gene, and nuclear matrix-like attachment sites flank the RBF element [Lauber et al. (1997) J. Biol. Chem. 272, 24657-24665]. In this paper, electrophoretic mobility shift assays (EMSAs) and S1 nuclease treatment are used to demonstrate that the RBF-maltose binding protei (MBP) fusion protein binds to single-stranded DNA of its element. Only the N-terminal domain of RBF binds the RBF DNA element as demonstrated by southwestern blot analyses, and by competition EMSAs between RBF-MBP and the N-terminal domain. Mass spectrometric analysis of the C-terminal domain of RBF demonstrates its potential to form noncovalent protein-protein interactions via a potential leucine-isoleucine zipperlike structure, suggesting a homo- and/or possible heterodimer structure in solution. These data support that the nuclear matrix binding site (acceptor site) for PR in the c-myc gene promoter is composed of RBF dimers bound to a specific single-stranded DNA element. The dimers of RBF are generated by C-terminal leucine zipper and the DNA binding occurs at the N-terminal parallel beta-sheet DNA binding motif. This complex is flanked by nuclear matrix attachment sites.